DNA-DNA hybridization of single-copy DNA sequences

Abstract

This chapter is reviews the basic principles of hybridization and the kinetics of reassociation. It provides an overview that summarizes and compares different techniques used in single-copy hybridization. The chapter then examines different estimates of distance derived from melting curves. Native DNA is isolated and purified to remove RNA and protein. Long-stranded DNA is then sheared to short fragments to permit the separation of repetitive and single-copy DNA and to reduce viscosity and gel formation. The chapter also explains the kinetics of reassociation. Rates of reassociation of DNA are influenced by several factors including genome complexity, DNA concentration, fragment size, reassociation temperature, and cation concentration. The complexity of the sheared genome is the length in base pairs (bp) of the longest nonrepeating sequence that is produced by splicing together fragments in the population. The rate of reassociation is inversely proportional to the complexity of the genome. The hybridization of tracer DNA with driver DNA is also overviewed in the chapter.

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